Proper formation of the kidney into a fully functional organ requires the integration of multiple signaling events. During the reiterative process of kidney tubule induction, a delicate balance is struck between signals that maintain a progenitor cell population and those that lead to differentiation. A tip in the balance toward either side has catastrophic consequences. A failure to convert progenitor cells into differentiated epithelia can result in either hypoplasia or progenitor cell-derived cancers, such as Wilms'tumors. Alternatively, if the proper number of progenitor cells is not maintained, they will be depleted prematurely, preventing further nephrogenesis and once again resulting in hypoplasia. Currently, we know little about the molecular mechanisms that control the balance between progenitors and differentiated tubular cells during tissue homeostasis or how these signals are regulated during normal development of the kidney. One of the central dogmas in kidney progenitor cell biology has been that canonical Wnt signaling must be repressed in the progenitor cells in order to keep them undifferentiated. However, we have recently discovered that not only is the canonical Wnt signaling pathway active in progenitors, it may in fact have higher levels of activity in this cell type than in the rest of the metanephric mesenchyme. Specifically, we have found that Wnt9b activates expression of a distinct set of target genes specifically within the progenitor cells and that this occurs through a b-catenin dependent mechanism. We hypothesize that high levels of canonical Wnt signaling activity in progenitor cells are promoted by the canonical Wnt agonists, R-spondin1 and 3, and that the expression of these genes in progenitor cells is dependent on the activity of the transcription factor Six2. However, several questions still remain: Why do some kidney mesenchymal cells differentiate into tubules, while others are maintained in a progenitor pool? What molecules regulate the choice between these alternative cellular states? Does Wnt9b signaling play a role in both populations? And which effectors modulate their different responses? The experiments outlined in this proposal are designed to test our hypotheses, as well as to enhance our understanding of the factors that regulate Wnt9b signaling and progenitor cell biology.PUBLIC HEALTH RELEVANCE: Unlike some organs, such as the pancreas, the functional unit of the kidney is not a single cell but instead a collection of cells organized into a tubule. The future of bioengineering tissue for organ replacement, at least for the kidney and other similar organs like the lung, thus relies not only on our ability to identify precursor or "stem" cells but also in our ability to convert these cells in functional tubules. We are interested in understanding the molecular and cellular processes that are required to convert individual cells into tubules.